Mycoplasmas cause a range of acute and chronic infections in humans. Mycoplasma genitalium is implicated as an emerging etiological agent of sexually transmitted and respiratory diseases and arthritides in humans and has been isolated from urethral, airway and synovial specimens. M. genitalium adheres to and colonizes host cell surfaces and establishes intracellular residence by mechanisms that remain poorly defined. However, we have shown that the distinct tip attachment organelle of M. genitalium is comprised of adhesins and adherence-related proteins, which are similar but not identical to those of Mycoplasma pneumoniae. Overall, very little is known about the biology and pathogenicity of M. genitalium. Our long-term goal is to delineate the mechanisms of cytadherence and invasion of M. genitalium and understand the dynamics of the M. genitalium-host interplay. In the Progress Report we describe ongoing studies of MG218 (tip organelle, cytadherence-accessory protein) and its associated operon and identify previously undetected, yet seemingly important M. genitalium mucin (Mn) binding proteins (MnBPs). Mucus, which contains Mn in high quantities, is a common substance in genitourinary, gastrointestinal and respiratory tracts, and the occurrence of MnBP in M. genitalium provides a novel virulence determinant. Furthermore, we have observed differential gene expression by M. genitalium in response to host cells and their products. These research developments suggest that mycoplasma-host interactions are more intricate than previously envisioned, and our primary goal is to delineate potential pathogenic mechanisms with the following specific aims: 1.Characterize genes and gene products of the mg218-associated operon as they influence mycoplasma adherence and invasion; 2. Investigate the role of M. genitalium MnBPs in pathogenesis; and 3. Identify mycoplasma genes participating in parasitism using DNA microarrays and real-time RT-PCR. We believe that the proposed work is innovative and capitalizes on the latest developments in genomics, particularly global analysis of gene expression in DNA microarray. It also takes advantage of our recently developed methods to disrupt genes in mycoplasmas through homologous recombination. It is our expectation that the proposed studies will significantly increase our understanding of mycoplasma pathogenicity and advance new therapeutic strategies to control mycoplasma infections and disease progression.